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Significance The paper proposes and demonstrates an approach for collecting solution NMR spectra from RNAs with high sensitivity, based on the chemical exchanges that labile hydrogens in the imino groups of nucleic acids can undergo with hyperpolarized water. This can lead to hundred-fold sensitivity enhancements for these diagnostic resonances, providing a tool to monitor complex RNA-based interconversions via 1-dimensional or 2-dimensional spectroscopy. The potential of this technology is demonstrated here by following the folding of the guanine-sensing riboswitch aptamer domain upon injection of hypoxanthine. This is a refolding process occurring within ∼1-s timescales, whose intermediates can be followed with atomic-level resolution under physiological temperatures and conditions, thanks to the enhancements brought by hyperpolarized NMR.

A 300-fold enhancement of imino nucleic acid resonances by hyperpolarized water provides a new window for probing RNA refolding by 1D and 2D NMR